The objectives of this research proposal are to examine the cellular controls of the transport proteins for the normal absorption of vitamin B12 (cobalamin) from the gastrointestinal tract. This information will be used to improve our understanding of the regulation of selected vitamin and mineral absorption as well as the pathology associated with malfunction of the regulatory mechanisms. We will study the synthesis and secretion of intrinsic factor (IF) from the guinea pig parietal cell using in vivo and in vitro (isolated parietal cells) models. Cellular events will be ascertained by making temporal comparisons between the microanatomical distribution of IF, as defined by ultrastructural immunocytochemistry, and the secretion of acid and IF into gastric juice, measured by RIA. These comparisons will be made before and after 1) IF secretagogues are given, 2) microtubule function is inhibited, and 3) IF secretion is inhibited by histamine-H2 receptor blockade. In addition, the effect of aging on the cellular controls of IF synthesis and secretion will be evaluated in man and the guinea pig. The availability of homogeneously purified IF, canine IF-B12 receptor, specific anti-IF, and anti-receptor antisera, will allow the sequential ultrastructural immunocytochemical localization of the sites of IF-B12 binding which will help define whether intact IF is internalized into the enterocyte during B12 absorption. Using the same reagents, as well as a RIA for IF-B12 receptor, we will ascertain whether the quantity and distribution of receptor is influenced bymicrotubule function and/or intraluminal IF. We will examine what the role of tanscobalamin II(TCII) and TCII receptor might have in the transport of B12 through, and out, of the enterocyte. The presence of enterocyte TCII or TCII receptor will be studied with ultrastructural immunocytochemistry, using specific anti-TCII and anti-TCII receptor antisera. We will try to ascertain whether TCII found in ileal enterocytes is of ileal or extraintestinal origin.